Culturing of cells of various types has become a routine process in many laboratories. Cells are grown to harvest compounds, to test for various sensitivities to potentially toxic compounds and even to provide tissue for grafts. This work generally is a monoculture, i.e., cells of one type are grown in a suitable medium.
More recently, interest has developed in the co-culture of cells. Co-culture of cells involves growing one population of cells in the presence of another population of cells. Co-culture of cells is important for study of inflammation reactions, cell differentiation processes and blood brain permeability studies.
Representative literature reports related to cell co-culture include: Magnum et al., In Vitro Cell Dev. Biol. 26:1135-1143 (Dec. 1990), "Co-Culture of Primary Pulmonary Cells to Model Alveolar Injury and Translocation of Protein"; Madara et al., J. Tissue Cult. Method, 14:209-216, (1992), "A Simple Approach to Electrical Parameters of Cultured Epithelial Monolayers: Use in Assessing Neutrophil-Epithelial Interactions"; Miller et al., J. Tissue Cult. Method, 14:217-224, "Application of Cultured Endothelial Cells of the Brain Microvasculature in the Study of the Blood-Brain Barrier"; and Science, 266:564-565 (1995), "Finding Clues About How Embryo Structures Form." The above referenced articles are cited to provide background regarding the in vitro study of interaction between one cell type and another.
Miller et al., cited above, describes culturing of cells on solid plastic surfaces and filters or membrane inserts. Miller et al. reports that bovine brain endothelial cells (BBEC) cultured on filters or membrane inserts provides an advantage over BBEC cultured on solid plastic surfaces. This advantage is that cell polarity with respect to metabolism or receptor distribution can be examined. Miller et al. further states that BBEC culture on filters or membrane inserts are required for determining the trans cellular transport or permeability of a compound across the cellular monolayer.
The above referenced Science article describes work done on kidney development. The article reports Mesenchymal cells co-cultured with cells producing Wnt-1 protein differentiate into kidney structures, including nephron tubular and glomular tissue and reports that this effect is not seen with control cells.
In response to the developing need for devices and equipment to co-culture cells, a co-culture system is disclosed in the commonly assigned U.S. patent application Ser. No. to Mussi et al. 08/124,415, now U.S. Pat. No. 5,409,829. The disclosure provides a complete self-contained system for preparing a co-culture of cells.
U.S. Pat. No. 5,026,649 to Lyman et al. discloses an insert device that can be utilized to culture and co-culture cells.
U.S. Pat. No. 4,871,674 to Matsui et al. discloses an insert for culturing cells having a porous membrane forming the bottom of a cylinder. The cylinder additionally has provisions for being suspended in a well.
Both the Lyman et al. disclosure and the Matsui et al. disclosure can be used to culture cells on a membrane, but neither is well suited for growing populations of cells on opposite sides of a membrane. The cell culture system disclosed in U.S. patent application Ser. No. 08/124,415 is well suited to culture cells on both sides of a membrane, but requires a series of manipulations that may be time consuming for screening studies where multiple co-cultures are being developed.
Additionally, there are reports in the literature suggesting that it is desirable to substantially eliminate air bubbles on the underside of the membrane during cell growth on the top side of the membrane, because uniform cell growth on the membrane top surface is inhibited on membrane regions having bubbles on the underside. Removal of bubbles from the underside of the membrane may prove difficult with the referenced devices.
In view of the increasing interest in co-culture of cells, there is a need for a simple to use apparatus for co-culture of cells that enables the practitioner to substantially avoid air bubbles on the underside of the membrane. Such a device and a method for it use is described herein below.